Spinal cord injury-induced immune deficiency syndrome enhances infection susceptibility dependent on lesion level

Abstract

Pneumonia is the leading cause of death after acute spinal cord injury and is associated with poor neurological outcome. In contrast to the current understanding, attributing enhanced infection susceptibility solely to the patient's environment and motor dysfunction, we investigate whether a secondary functional neurogenic immune deficiency (spinal cord injury-induced immune deficiency syndrome, SCI-IDS) may account for the enhanced infection susceptibility. We applied a clinically relevant model of experimental induced pneumonia to investigate whether the systemic SCI-IDS is functional sufficient to cause pneumonia dependent on spinal cord injury lesion level and investigated whether findings are mirrored in a large prospective cohort study after human spinal cord injury. In a mouse model of inducible pneumonia, high thoracic lesions that interrupt sympathetic innervation to major immune organs, but not low thoracic lesions, significantly increased bacterial load in lungs. The ability to clear the bacterial load from the lung remained preserved in sham animals. Propagated immune susceptibility depended on injury of central pre-ganglionic but not peripheral postganglionic sympathetic innervation to the spleen. Thoracic spinal cord injury level was confirmed as an independent increased risk factor of pneumonia in patients after motor complete spinal cord injury (odds ratio = 1.35, P < 0.001) independently from mechanical ventilation and preserved sensory function by multiple regression analysis. We present evidence that spinal cord injury directly causes increased risk for bacterial infection in mice as well as in patients. Besides obvious motor and sensory paralysis, spinal cord injury also induces a functional SCI-IDS ('immune paralysis'), sufficient to propagate clinically relevant infection in an injury level dependent manner.

Keywords: mechanisms; myelopathy; neuroinflammation; rehabilitation; spinal cord injury.

Infections are the main cause of death after spinal cord injury (SCI). Brommer et al. reveal that an SCI-induced immune deficiency syndrome is responsible for the enhanced infection susceptibility in SCI patients and a mouse model. Lesion level determines the extent of this ‘immune paralysis’.

Figure 1

Human and controlled experimental SCI-associated pneumonia (SCI-AP). (A) Typical SCI-AP occurs with an onset from Day 2–7 (Berlly and Shem, 2007) in humans and is caused by community-acquired pathogens, prevailingly S. pneumonia (white arrows: pneumonic infiltrates). SCI-AP is antedated by onset of SCI-IDS beginning within the first 24 h (Riegger et al., 2007, 2009). (B) Experimental design to investigate whether SCI lesion level enhances the susceptibility to infection after SCI. Mice were treated with antibiotics starting 1 day before surgery until the day before infection to prevent uncontrolled spontaneous infections. Animals received either a laminectomy only (control group) or a transection injury at thoracic level 3 or 9 (Th3, Th9). Animals were inoculated with 500 cfu of S. pneumoniae on Day 3 after surgery by a lung intubation. 24 h after infection, the animals were killed and the organs were harvested and processed. (C) To investigate the origin of the maladaptive signalling after SCI a preceding peripheral sympathetic deafferentation of the spleen (and a laparotomy as appropriate control) occurred 12 days before SCI. Here we investigate whether SCI-IDS is functional relevant and able to propagate the susceptibility for bacterial infections in a lesion dependent manner (neurogenic implication).

Figure 2

Experimental SCI with differential lesion levels (Th3 versus Th9) of the sympathetic preganglionic neurons. (A) The CNS controls the immune system through the autonomous nervous system. SCI specifically disconnects sympathetic preganglionic neurons located in the thoracolumbar spinal cord, which control systemic immune function by postganglionic noradrenergic projections to secondary lymphoid tissue including the spleen. Inhibition of spinally generated SNA and reflex excitation through sympathetic preganglionic neurons by baroreceptors and other supraspinal inhibitory systems (bulbo-spinal tracts) is lost after complete SCI above mid-thoracic level (‘isolated sympathetic spinal cord’ by ‘sympathetic decentralization’). Episodes of excessive SNA correlate with injury severity and lesion level. Lesion level dependent injury of sympathetic preganglionic neurons localized to the inter-mediolateral columns (IML) and funciculus lateralis (FL) in the thoracic rodent spinal cord is assured by a dorsal > 4/5 transection SCI. (B–D) Severity, consistency and SCI lesion level were confirmed by MRI 1 day before infection. Animals with spared ventral tissue bridges were excluded. Representative T₂ MRI images, illustrating a spinal cord lesion at the respective SCI level Th3 (B) and Th9 (C) (white arrows) compared with the uninjured spinal cord after sham surgery (D).

Figure 3

Differential sympathetic deafferentation level determines infection susceptibility. (A) We assessed the infection susceptibility after Th3-SCI and Th9-SCI compared to sham operated animals differing in terms of sympathetic decentralization. (B) The elevated bacterial load in the lung 24 h after inoculation demonstrated an increased susceptibility for bacterial infection in SCI animals (11 of 17, 65 %) compared to sham animals (1 of 7, 14%). Th3 level SCI (eight of nine, 89%) triggered a significantly elevated susceptibility compared to Th9 level SCI (three of eight, 38%). Besides obvious motor and sensory paralysis SCI induces a functional SCI-IDS (‘immune paralysis’) causally relevant for clinical prevalent infections. Lines indicate median. Kruskal-Wallis analysis with Dunn’s post-test, *P < 0.05, **P < 0.01. (C) Haematoxylin and eosin staining of lung tissue from a Th3-injured mouse within the first 24 h after infection, demonstrating histopathological hallmarks of human pneumonia such as massive infiltration of immune cells and oedema formation. (D) Tissue of uninjured, uninfected mouse for comparison.

Figure 4

Spleen atrophy is only caused after high thoracic but not after low thoracic SCI. To investigate the relevance of efferent sympathetic innervation of secondary lymphoid organs we focused on the spleen as the largest secondary lymphoid organ and investigated spleen atrophy as a macroscopic marker associated with systemic immune dysfunction. Shown are representative images of spleens derived from sham animals compared with mice after Th3 and Th9 SCI. (A) Spleen atrophy is induced dependent on sympathetic decentralization by Th3 SCI but not by Th9 SCI with spared efferent sympathetic innervation. (B) Data are presented as box plots with whiskers showing minimum to maximum. Th3- and Th9-level SCI mice do not differ in terms of vagus nerve innervation, which is derived from supra-lesional brainstem regions and is reported to also exert immune-modulatory effects. One-way ANOVA analysis with Bonferroni post tests, n as in Fig. 3; *P < 0.05, **P < 0.01.

Figure 5

Functional SCI-IDS depends on central (preganglionic) lesion site and is not caused by postganglionic sympathetic deafferentation. (A) We investigated whether deafferentation of sympathetic efferents per se causes enhanced immune susceptibility or whether functional immune suppressive effects depend on its injury site [central (preganglionic) versus peripheral (postganglionic sympathetic injury); vertical grey dotted line demarcates the border between pre- and postganglionic sympathetic innervation, localized at the ganglion coeliacum]. As shown above, a high-thoracic preganglionic injury causes functional SCI-IDS (Fig. 3) and spleen atrophy (Fig. 4). However, postganglionic deafferentation (peripheral injury to the splenic nerve prior to a Th9-SCI, which itself does not cause SCI-IDS; Th9-SD) does not enhance bacterial load comparing with intact peripheral spleen innervation (Th9-lap) (B) and does not affect spleen size (C). Thus, postganglionic sympathetic deafferentation of large lymphoid organs does not propagate infection susceptibility. Bacterial load in lungs are displayed on a logarithmic scale with lines indicating medians. Spleen lengths are displayed as box plots with whiskers reaching from minimum to maximum. SD = spleen deafferentation; Lap = laparotomy (sham surgery).

Figure 6

Maladaptive sympathetic signalling triggering functional SCI-IDS after high thoracic SCI originates from below the lesion site. (A) Given that enhanced infection susceptibility (functional SCI-IDS) is induced by central (preganglionic) and high (Th3-SCI) injury only, we analysed whether the underlying mechanism is dependent on disinhibited spinally generated SNA originating distal to the lesion site with unrestricted access into the spleen. Therefore, we analysed the effect of blocking SNA signalling to the spleen by deafferentation of the splenic nerve. (B) Peripheral, postganglionic sympathetic deafferentation of the spleen prior Th3-SCI (Th3-SD, ‘spleen shielding’ from SNA) decreases the bacterial load in the lung 24 h after infection (improved host defence) compared to sham/laparotomy operated Th3-SCI animals (Th3-lap) (Mann-Whitney test, Th3-SD versus Th3-lap, P = 0.06). (C) Congruently, it attenuates spleen size atrophy after Th3-SCI (t-test, Th3-SD versus Th3-lap P = 0.06). The fact that compensatory signals cannot reach the ganglion coeliacum and the spleen in case of Th3 SCI, supports the hypothesis that the origin of maladaptive sympathetic signalling is below the injury (A). Bacterial loads in lungs are displayed on a logarithmic scale with lines indicating medians. Spleen lengths are displayed as box plots with whiskers reaching from minimum to maximum. SD = spleen deafferentation; Lap = laparotomy (sham surgery). Vertical grey dotted line demarcates the border between pre- and postganglionic sympathetic innervation localized at the ganglion coeliacum.

Figure 7

Thoracic lesion level and severity after human SCI are independently associated with enhanced ‘neurogenic’ risk of SCI-associated pneumonia (SCI-AP). (A) Sympathetic innervation (preganglionic, postganglionic) of immunological relevant organs in humans. SCI rostral to Th5 (Th1–Th4) will result in preganglionic injury to sympathetic efferents whereas Th5–Th8 SCI spares some and SCI Th9–Th12 most sympathetic efferents to the ganglion coeliacum. (B) We compared motor complete (AIS A and B patients, grey bars, given the excellent correspondence between somatic and complete sympathetic lesions in this population) with motor incomplete patients (AIS C and D, white bars) with enhanced sparing of sympathetic outflow. Relative frequency of pneumonia is shown in motor complete versus motor incomplete SCI patients, categorized as high (Th1–Th4), mid (Th5–Th8) or low (Th9–Th12) thoracic injury level. Within the group of Th1–Th4 patients, pneumonia rates are significantly different between motor complete and incomplete patients, whereas no differences between the groups were detectable in lower thoracic levels. Error bars represent 95% CI. Differences within the neurological level categories were assessed with the chi-square test. Patient numbers: motor complete Th1–Th4, n = 317, Th5–Th8 n = 271, Th9–Th12 n = 451; motor incomplete Th1–Th4, n = 41, Th5–Th8 n = 34, Th9–Th12 n = 107.

Figure 8

Neurogenic pathophysiology and targets of maladaptive sympathetic signalling perpetuating functional SCI-IDS after SCI. Under physiological conditions excitatory sympathetic signals are controlled by supraspinal inhibition. After SCI, loss of supraspinal control leads to spinally generated sympathetic nerve activity (SNA). SNA originates below SCI injury and leads to maladaptive efferent sympathetic activity signalling to the spleen. It is able to culminate in large excitatory spinal sympathetic reflexes. Spinal sympathetic reflexes occur in analogy to pathological Babinksi motor reflexes, caused by a loss of supraspinal (bulbospinal) tonic inhibition (‘Sympathico-Babinski’). Disinhibited spinally generated sympathetic nerve activity after high thoracic (Th3) SCI leads to ad hoc induction of excitatory SNA burst entering the splenic nerve (Dembowsky et al., 1980; Taylor and Schramm, 1987) associated with increased levels of norepinephrine (NE) in the spleen, which in turn causes apoptosis of immune cells in the spleen (Lucin et al., 2007, 2009). This results in a decrease of spleen size and an elevated susceptibility to infection. Blocking SNA signaling to the spleen (‘shielding’) by preceding peripheral denervation of the splenic nerve ameliorates functional SCI-IDS and bacterial load after Th3-SCI. Moreover, here we identify the peripheral splenic nerve as a target for immunomodulation after SCI in order to restore impaired host defense after SCI (SCI-IDS) by blocking SNA to the spleen. Spleen shielding may be an interventional strategy to ameliorate functional SCI-IDS to prevent infections in patients at risk.